Cloning, expression, and mapping of the Staphylococcus aureus alpha-hemolysin determinant in Escherichia coli K-12.

A fragment of Staphylococcus aureus DNA encoding the alpha-hemolysin determinant was cloned from strain Wood 46 by inserting Sau3A-generated genomic DNA fragments between the BamHI sites of the lambda replacement vector L47.1. Phages expressing alpha-hemolysin were detected by overlaying plaques formed from several thousand independent recombinant phage with erythrocytes and looking for zones of hemolysis. One phage expressing alpha-hemolysin was purified and named lambda w alpha 3. This was subsequently shown to contain a 10.2-kilobase pair insert of S. aureus DNA. A 7.6-kilobase pair HindIII fragment encoding the alpha-hemolysin was subcloned from lambda w alpha 3 into the plasmid vector pACYC184 to form the hybrid plasmid pDU1148. Escherichia coli K-12 cells harboring pDU1148 synthesized a low level of alpha-hemolysin which remained associated with the cells and was not secreted into culture supernatants. When the same strain was stabbed onto blood agar plates, no zones of hemolysis were detected after overnight growth at 37 degrees C but hemolysis developed if the plates were left at room temperature for 48 h. By introducing specific deletions or Tn5 insertions into plasmid pDU1148, the alpha-hemolysin gene was mapped to a region within a 3.3-kilobase pair EcoRI-HindIII fragment which was subcloned onto the vector plasmid pBR322. A specific enzyme-linked immunosorbent assay with peroxidase-labeled rabbit anti-alpha-hemolysin antibodies was used to measure the levels of alpha-hemolysin antigen expressed in E. coli K-12 cells harboring pDU1148 or a variety of pDU1148::Tn5 and pDU1148 deletion mutants.     

CD163: a specific marker of macrophages in paraffin-embedded tissue samples

Paraffin-section immunohistochemical analysis was performed using a monoclonal antibody against CD163 to evaluate the antibody's usefulness in identifying cells of monocyte/macrophage lineage in normal and neoplastic conditions. Normal human tissue samples and samples from 211 hematopoietic disorders and 115 nonhematopoietic neoplasms were examined. The distribution of KP1 and PG-M1, monoclonal antibodies to the macrophage-associated CD68 antigen, also were evaluated for comparison. CD163 immunoreactivity was observed in resident macrophages of all normal tissue samples except splenic white pulp macrophages and germinal center tingible body macrophages. Among hematopoietic disorders and nonhematopoietic neoplasms, CD163 expression was restricted largely to cases of chronic myelomonocytic leukemia, histiocytic sarcoma, sinus histiocytosis with massive lymphadenopathy, and littoral cell angioma. Acute myeloid leukemias (AMLs) with monocytic differentiation were CD163- with the exception of 1 case of acute monoblastic leukemia. Most myeloid sarcomas also were CD163-. Compared with the CD68 antibodies, CD163 demonstrated greater specificity as a marker of disorders of monocyte/macrophage origin. However, immunohistochemical evaluation of CD163 expression does not seem to be a sensitive means of determining monocytic differentiation in AMLs in paraffin sections or establishing a diagnosis of myeloid sarcoma.     

Contraceptive use among South Asian women attending general practices in southwest London.

A cross-sectional survey of contraceptive practices was conducted among 180 South Asian women aged 16 to 50 years, attending inner-city general practices. Overall prevalence of contraceptive use was 63% and ranged from 70% in South Asian teenagers, to only 50% in women over 30 who had completed their family. Thirteen per cent of women had had a termination of pregnancy Although contraceptive use in this group is increasing, it has not yet reached national levels.     

Subcutaneous phaeohyphomycosis caused by Cladophialophora bantiana

Primary subcutaneous phaeohyphomycosis can rarely be caused by Cladophialophora bantiana, and we present the histologic and culture findings of such a case. A 32-year-old African American woman with systemic lupus erythematosus presented with a 2-year history of multiple, recurrent, tender, and ulcerated skin nodules with purulent drainage on her upper back. Histologic sections of the excision demonstrated features of phaeohyphomycosis. Culture findings were characteristic of C bantiana. Of interest, at age 10 she had sustained traumatic implantation of wood splinters into this area during a tornado, yet clinical symptoms of a subcutaneous infection did not manifest until she developed lupus erythematosus at age 27. Our case highlights the role of trauma and immunosuppression in the pathogenesis of subcutaneous phaeohyphomycosis.     

Utility of fine-needle aspiration in the diagnosis of primary osteosarcoma

Fine-needle aspiration (FNA) is a reliable, safe, and cost-effective procedure with a well-established role in the diagnosis of various solid tissue neoplasms. The role of FNA in the diagnosis of primary bone tumors, including osteosarcoma (OGS), is controversial and has yet to be established. We reviewed our experience with the use of FNA as a diagnostic technique over the past 8 yr at our institution. Diagnosis was conclusive in 26 (65%) of 40 patients, 18 of whom went to neoadjuvant therapy and/or resection based solely on the FNA interpretation of either "high grade sarcoma" or "osteosarcoma." Of the remaining 14 (25%) patients, 12 had inconclusive diagnosis and two (5%) were false-negatives. An inconclusive diagnosis was most likely to be an inadequate or paucicellular aspirate, seen in six (15%) patients. An additional six patients had variants of osteosarcoma (four chondroid, one "giant cell rich," one parosteal) that made definitive diagnosis impossible. The two that were incorrectly classified were diagnosed as fracture callus and plasmacytoma. FNA is an accurate and cost-effective tool for the initial diagnosis of primary osteosarcoma with a sensitivity of 65% and accuracy of 95%. Inconclusive diagnoses are likely to be due to insufficient sample cellularity or the presence of OGS variant. In our experience, FNA is sufficient to provide the diagnosis of OGS prior to definitive treatment when interpreted in conjunction with imaging studies and clinical findings. In those cases where FNA fails to yield a diagnostic sample, a traditional biopsy can be performed.     

Stroke-prone rats exhibit prolonged behavioral deficits without increased brain injury: an indication of disrupted post-stroke brain recovery of function

Stroke-prone rat strains exhibit an increased stroke risk and sensitivity, and reduced endogenous mechanisms of ischemic brain tolerance. This experiment provides a comparative, serial evaluation of neurological deficits and brain injury following middle cerebral artery occlusion/permanent focal stroke in this high-risk strain. Stroke-prone spontaneously hypertensive (SHR-SP), spontaneously hypertensive (SHR) and Wistar Kyoto (WKY) rats were evaluated over 28 days using magnetic resonance imaging (MRI), histopathology, and neurobehavioral testing. T2- and diffusion weighted-MRI was performed after 1, 10 and 28 days to measure the degree of stroke-induced brain injury. Normotensive WKY rats receiving the same stroke and other SHR-SP rats receiving sham surgery were used for control comparisons. Functional deficits were scored after 1, 4, 11, 18 and 28 days. The degree of brain infarction/injury was practically identical in hypertensive and stroke-prone rats. WKY rats exhibited significantly smaller infarcts (P<0.05), with neurological function recovering quickly to normal by day 11 in this strain. Functional deficits persisted longer in hypertensive rats, with function recovering to normal by day 18 (P<0.05). Functional deficits in SHR-SP rats persisted the longest, and were observed even after 28 days (P<0.05). This increased and prolonged neurologic dysfunction exhibited by SHR-SP compared to SHR rats, while exhibiting practically identical degrees of brain injury/infarction, reflects the increased stroke risk and sensitivity of this strain and suggests a reduced SHR-SP brain plasticity following injury. Therefore, the stroke-prone rat provides an enhanced and prolonged functional deficit model that can be used to elucidate those mechanisms/novel targets critical to longitudinal neurobehavioral recovery post-stroke.     

Determinants of cockroach and mouse exposure and associations with asthma in families and elderly individuals living in New York City public housing.

BACKGROUND: Asthma prevalence is high in the inner city, and morbidity has been associated with cockroach and mouse allergens. OBJECTIVE: To characterize the relationships among pests, allergens, pesticides, and asthma in New York City public housing. METHODS: In 324 apartments, dust samples collected from beds and kitchens were analyzed for cockroach (Bla g 2) and mouse (mouse urinary protein [MUP]) allergens, pest populations were monitored, and residents were interviewed about home characteristics and asthma symptoms. RESULTS: Cockroaches were found in 77% of the apartments, and evidence of mice was found in 13%. Allergens and pesticide use were associated with pest infestation, and 15% of residents reported using illegal pesticides. The percentage of apartments with high allergen levels varied significantly by building (Bla g 2: P = .002; MUP: P = .03), as did the percentage of apartments with cockroaches (P = .002) and daily mouse sightings (P = .02). Thirty-seven percent of the apartments had at least 1 resident with physician-diagnosed asthma. In family buildings, apartments with high Bla g 2 levels had 1.7 times greater odds of having an asthmatic resident (95% confidence interval, 1.2-2.3). In senior citizen buildings, apartments with high MUP levels had 6.6 times greater odds of having an asthmatic resident (95% confidence interval, 1.4-31.7), controlling for smoking and other potential confounders. CONCLUSIONS: Previous studies have identified home characteristics associated with the presence of cockroaches and mice, but the present findings suggest that building-level characteristics can affect high pest exposure. Furthermore, the high asthma prevalence in residents and the use of illegal pesticides highlight the need for safe and effective building-wide pest control strategies.